Cardiac failure occurs in patients with diabetes mellitus inthe absence of coronary vascular disease. The decreased ability of the diabetic heart to contract may result from specific quantitative and qualitative changes which occur in cardiac proteins. We have found previously that one such change is represented by the diabetes-induced increased predominance of myosin V3. The normal rat ventricle contains the myosin isoenzymes V1, V2, and V3 and in the normal heart myosin V1 represents 70% of all myosin. In contrast in diabetic hearts 68% of all myosin constitutes myosin v3. The increased myosin V3 predominance is accompanied by a 45% decrease in Ca++ activated myosin ATPase activity. A close correlation between the activity of this enzyme and the velocity of shortening of cardiac muscle has been established. Myosin V1 contains most likely two identical myosin heavy chain species (MHC-alpha) which are different from the two MHC-beta species contained in myosin V3. Data recently obtained by us indicate that the increased myosin V3 level in diabetic hearts is related to increased levels of MHC-beta. We will investigate during this grant period if the level of MHC-beta mRNA and MHC-alpha mRNA themselves change in the diabetic heart. For the precise quantitation of the respective mRNAs specific cDNA probes will be obtained. In addition we will determine the influence which diabetes mellitus has on the level of other specific cardiac mRNAs which do not code for MHCs. For this purpose mRNAs will be translated in a modified rabbit reticulocyte lysate system, translational products will be separated by two-dimensional gel electrophoresis and quantitated by digital matrix photometry. The time course of changes in the level of these mRNAs after insulin administration will also be determined. To identify the specific pathophysiological alterations of the diabetic milieu which mediate the increase in myosin V3 is one of the aims of this grant application. For this purpose glucose and fatty acid consumption will be determined in myocytes obtained from normal, diabetic and diabetic rats treated with insulin and other compounds. Changes in substrate consumption will be correlated with the relative synthesis rate of MHC-alpha and MHC-beta and with mHC mRNA levels.
